A 3-dimensional microfluidic platform for modeling human extravillous trophoblast invasion and toxicological screening

Hemochorial placentation is characterized by the development of trophoblast cells specialized to interact with the uterine vascular bed. We utilized trophoblast stem (TS) cell and mutant rat models to investigate regulatory mechanisms controlling trophoblast cell development. TS cell differentiation was characterized by acquisition of transcript signatures indicative of an endothelial cell-like phenotype, which was highlighted by the expression of anticoagulation factors including tissue factor pathway inhibitor (TFPI). TFPI localized to invasive endovascular trophoblast cells of the rat placentation site. Disruption of TFPI in rat TS cells interfered with development of the endothelial cell-like endovascular trophoblast cell phenotype. Similarly, TFPI was expressed in human invasive/extravillous trophoblast (EVT) cells situated within first-trimester human placental tissues and following differentiation of human TS cells. TFPI was required for human TS cell differentiation to EVT cells. We next investigated the physiological relevance of TFPI at the placentation site. Genome-edited global TFPI loss-of-function rat models revealed critical roles for TFPI in embryonic development, resulting in homogeneous midgestation lethality prohibiting analysis of the role of TFPI as a regulator of the late-gestation wave of intrauterine trophoblast cell invasion. In vivo trophoblast-specific TFPI knockdown was compatible with pregnancy but had profound effects at the uterine–placental interface, including restriction of the depth of intrauterine trophoblast cell invasion while leading to the accumulation of natural killer cells and increased fibrin deposition. Collectively, the experimentation implicates TFPI as a conserved regulator of invasive/EVT cell development, uterine spiral artery remodeling, and hemostasis at the maternal–fetal interface.

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3D Bioprinted GelMA Based Models for the Study of Trophoblast Cell Invasion

Scientific Reports ◽

10.1038/s41598-019-55052-7 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 4

Author(s):

Houzhu Ding ◽

Nicholas P. Illsley ◽

Robert C. Chang

Keyword(s):

Cell Invasion ◽

Functional Limitations ◽

Cell Movement ◽

Trophoblast Cell ◽

Trophoblast Invasion ◽

Ring Model ◽

Cell Front ◽

A Cell ◽

Practical Tool ◽

And Migration

AbstractBioprinting is an emerging and promising technique for fabricating 3D cell-laden constructs for various biomedical applications. In this paper, we employed 3D bioprinted GelMA-based models to investigate the trophoblast cell invasion phenomenon, enabling studies of key placental functions. Initially, a set of optimized material and process parameters including GelMA concentration, UV crosslinking time and printing configuration were identified by systematic, parametric study. Following this, a multiple-ring model (2D multi-ring model) was tested with the HTR-8/SVneo trophoblast cell line to measure cell movement under the influence of EGF (chemoattractant) gradients. In the multi-ring model, the cell front used as a cell invasion indicator moves at a rate of 85 ± 33 µm/day with an EGF gradient of 16 µM. However, the rate was dramatically reduced to 13 ± 5 µm/day, when the multi-ring model was covered with a GelMA layer to constrain cells within the 3D environment (3D multi-ring model). Due to the geometric and the functional limitations of multi-ring model, a multi-strip model (2D multi-strip model) was developed to investigate cell movement in the presence and absence of the EGF chemoattractant. The results show that in the absence of an overlying cell-free layer of GelMA, movement of the cell front shows no significant differences between control and EGF-stimulated rates, due to the combination of migration and proliferation at high cell density (6 × 106 cells/ml) near the GelMA surface. When the model was covered by a layer of GelMA (3D multi-strip model) and migration was excluded, EGF-stimulated cells showed an invasion rate of 21 ± 3 µm/day compared to the rate for unstimulated cells, of 5 ± 4 µm/day. The novel features described in this report advance the use of the 3D bioprinted placental model as a practical tool for not only measurement of trophoblast invasion but also the interaction of invading cells with other tissue elements.

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Interferon‐γ inhibits extravillous trophoblast cell invasion by a mechanism that involves both changes in apoptosis and protease levels

The FASEB Journal ◽

10.1096/fj.06-6616com ◽

2006 ◽

Vol 20 (14) ◽

pp. 2512-2518 ◽

Cited By ~ 89

Author(s):

Gendie E. Lash ◽

Harry A. Otun ◽

Barbara A. Innes ◽

Maureen Kirkley ◽

Leandro De Oliveira ◽

...

Keyword(s):

Cell Invasion ◽

Interferon Γ ◽

Trophoblast Cell ◽

Extravillous Trophoblast

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The depletion of MARVELD1 leads to placenta accreta via integrin β4-dependent trophoblast cell invasion

10.1101/106807 ◽

2017 ◽

Author(s):

Yue Chen ◽

Hui Zhang ◽

Fang Han ◽

Lei Yue ◽

Chunxiao Qiao ◽

...

Keyword(s):

Cell Invasion ◽

Placenta Accreta ◽

Wild Type Mouse ◽

Trophoblast Cell ◽

Trophoblast Invasion ◽

Trophoblast Cells ◽

Integrin Β4 ◽

Underlying Mechanisms

AstractThe mammalian placenta is a remarkable organ. It serves as the interface between the mother and the fetus. Proper invasion of trophoblast cells into the maternal decidua is required for a successful pregnancy. Previous studies have found that the adhesion molecule integrin β4 plays important roles during trophoblast cell invasion. Here, we found that the overall birth rate of the MARVELD1 knockout mouse is much lower than that of the wild-type mouse (P<0.001). In E18.5 MARVELD1 knockout mice, we observed an over-invasion of trophoblast cells, and indeed, the pregnant mice had a partial placenta accreta phenotype. The HTR8/SVneo cell line was used as an in vitro model to elucidate the underlying mechanisms of MARVELD1-mediated trophoblast invasion. We detected a diminished expression of integrin β4 upon the downregulation of MARVELD1 and enhanced migration and invasive abilities of trophoblast cells both in vivo and in vitro. The integrin β4 rescue assay also supported the results. In conclusion, this study found that MARVELD1 mediated the invasion of trophoblast cells via regulating the expression of integrin β4.

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Extravillous trophoblast cell invasion is promoted by the CD44–hyaluronic acid interaction

Placenta ◽

10.1016/j.placenta.2013.12.009 ◽

2014 ◽

Vol 35 (3) ◽

pp. 163-170 ◽

Cited By ~ 13

Author(s):

H. Takahashi ◽

T. Takizawa ◽

S. Matsubara ◽

A. Ohkuchi ◽

T. Kuwata ◽

...

Keyword(s):

Hyaluronic Acid ◽

Cell Invasion ◽

Trophoblast Cell ◽

Extravillous Trophoblast ◽

Acid Interaction

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Placenta-specific miRNA miR-520c modulates CD44-mediated extravillous trophoblast cell invasion

Placenta ◽

10.1016/j.placenta.2014.06.215 ◽

2014 ◽

Vol 35 (9) ◽

pp. A66-A67

Author(s):

Hironori Takahashi ◽

Akihide Ohkuchi ◽

Takami Takizawa ◽

Shigeki Matsubara ◽

Rie Usui ◽

...

Keyword(s):

Cell Invasion ◽

Trophoblast Cell ◽

Extravillous Trophoblast

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Delta-9-tetrahydrocannabinol inhibits invasion of HTR8/SVneo human extravillous trophoblast cells and negatively impacts mitochondrial function

Scientific Reports ◽

10.1038/s41598-021-83563-9 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

O’Llenecia S. Walker ◽

Harmeet Gurm ◽

Reeti Sharma ◽

Navkiran Verma ◽

Linda L. May ◽

...

Keyword(s):

Molecular Mechanisms ◽

Cannabinoid Receptors ◽

Mitochondrial Fission ◽

Placental Tissue ◽

Trophoblast Cell ◽

Extravillous Trophoblast ◽

Trophoblast Invasion ◽

Stress Markers ◽

Cellular Reactive Oxygen Species ◽

Important Health

AbstractPrenatal cannabis use is a significant problem and poses important health risks for the developing fetus. The molecular mechanisms underlying these changes are not fully elucidated but are thought to be attributed to delta-9-tetrahydrocannabinol (THC), the main bioactive constituent of cannabis. It has been reported that THC may target the mitochondria in several tissue types, including placental tissue and trophoblast cell lines, and alter their function. In the present study, in response to 48-h THC treatment of the human extravillous trophoblast cell line HTR8/SVneo, we demonstrate that cell proliferation and invasion are significantly reduced. We further demonstrate THC-treatment elevated levels of cellular reactive oxygen species and markers of lipid damage. This was accompanied by evidence of increased mitochondrial fission. We also observed increased expression of cellular stress markers, HSP70 and HSP60, following exposure to THC. These effects were coincident with reduced mitochondrial respiratory function and a decrease in mitochondrial membrane potential. Taken together, our results suggest that THC can induce mitochondrial dysfunction and reduce trophoblast invasion; outcomes that have been previously linked to poor placentation. We also demonstrate that these changes in HTR8/SVneo biology may be variably mediated by cannabinoid receptors CB1 and CB2.

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Bone Morphogenetic Protein 2 Increases Human Trophoblast Invasion by Up-Regulating Integrin Beta3

Journal of the Endocrine Society ◽

10.1210/jendso/bvab048.1520 ◽

2021 ◽

Vol 5 (Supplement_1) ◽

pp. A747-A748

Author(s):

Cuiping Hu ◽

Junhao Yan

Keyword(s):

Bone Morphogenetic Protein ◽

Trophoblast Cell ◽

Bone Morphogenetic Protein 2 ◽

Extravillous Trophoblast ◽

Trophoblast Invasion ◽

Trophoblast Cells ◽

Human Trophoblast ◽

Protein Levels ◽

Integrin Β3 ◽

Morphogenetic Protein

Abstract The adequate invasion of extravillous trophoblast cells (EVTs) is indispensable for the implantation of embryos and subsequent remodeling of uterine spiral arteries in early human gestation. Bone morphogenetic protein 2 (BMP2), which is abundantly expressed at the maternal-fetal interface, has been shown to promote the human EVT invasion process (1). Integrin switching (i.e., a switch from α6β4 to αvβ3) plays essential roles in cell-extracellular matrix adhesion and has been reported to influence EVT migration and invasion (2). Moreover, integrin β3 has been found to promote the adhesion, invasion, and migration abilities of embryonic trophoblasts (3). However, whether integrin β3 participates in BMP2 signaling and mediates BMP2-increased-human trophoblast invasion remains unknown. The purpose of our study was to explore the effects of BMP2 on integrin αvβ3 expression and the possible mediation role of integrin β3 in BMP2-regulated human trophoblast invasion. We used immortalized human trophoblast cell line (HTR8/SVneo) and primary human extravillous trophoblast cells (EVTs) isolated from first-trimester villi as study models. RT-qPCR and Western blot assay were respectively utilized to detect the messenger RNA and protein levels of intergrin αv and β3. The function of the target protein was studied by siRNA knockdown, and the trophoblast invasion ability was checked by Matrigel-coated transwell invasion assays. Our results demonstrated that the mRNA and protein levels of integrin β3, rather than integrin αv, were up-regulated after BMP2 treatment in HTR8/SVneo and primary EVT cells. Importantly, siRNA-mediated down-regulation of integrin β3 significantly inhibited basal and BMP2-induced HTR8/SVneo cell invasionas measured by transwell invasion assay. In conclusion, we findings support that BMP2 promotes human trophoblast cell invasion by up-regulating integrin β3 expression, benefiting the in-depth understanding of the pro-invasive effect of BMP2 on human trophoblasts during early pregnancy. Reference: (1) Hong-Jin Zhao et al., Cell Death Dis 2018;9:174. (2) Damsky, C.H. et al, Development 1994; 120, 3657-3666. (3) Dong-Mei He et al., Reproduction 2019;157:423-430.

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ASCL2 reciprocally controls key trophoblast lineage decisions during hemochorial placenta development

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2016517118 ◽

2021 ◽

Vol 118 (10) ◽

pp. e2016517118

Author(s):

Kaela M. Varberg ◽

Khursheed Iqbal ◽

Masanaga Muto ◽

Mikaela E. Simon ◽

Regan L. Scott ◽

...

Keyword(s):

Cell Differentiation ◽

Cell Invasion ◽

Cell Lineage ◽

Trophoblast Cell ◽

Extravillous Trophoblast ◽

Sequencing Analysis ◽

Spiral Artery ◽

Trophoblast Cells ◽

Junctional Zone ◽

Artery Remodeling

Invasive trophoblast cells are critical to spiral artery remodeling in hemochorial placentation. Insufficient trophoblast cell invasion and vascular remodeling can lead to pregnancy disorders including preeclampsia, preterm birth, and intrauterine growth restriction. Previous studies in mice identified achaete-scute homolog 2 (ASCL2) as essential to extraembryonic development. We hypothesized that ASCL2 is a critical and conserved regulator of invasive trophoblast cell lineage development. In contrast to the mouse, the rat possesses deep intrauterine trophoblast cell invasion and spiral artery remodeling similar to human placentation. In this study, we investigated invasive/extravillous trophoblast (EVT) cell differentiation using human trophoblast stem (TS) cells and a loss-of-function mutant Ascl2 rat model. ASCL2 transcripts are expressed in the EVT column and junctional zone, which represent tissue sources of invasive trophoblast progenitor cells within human and rat placentation sites, respectively. Differentiation of human TS cells into EVT cells resulted in significant up-regulation of ASCL2 and several other transcripts indicative of EVT cell differentiation. Disruption of ASCL2 impaired EVT cell differentiation, as indicated by cell morphology and transcript profiles. RNA sequencing analysis of ASCL2-deficient trophoblast cells identified both down-regulation of EVT cell-associated transcripts and up-regulation of syncytiotrophoblast-associated transcripts, indicative of dual activating and repressing functions. ASCL2 deficiency in the rat impacted placental morphogenesis, resulting in junctional zone dysgenesis and failed intrauterine trophoblast cell invasion. ASCL2 acts as a critical and conserved regulator of invasive trophoblast cell lineage development and a modulator of the syncytiotrophoblast lineage.

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